Summary: Developmental programs that control cell differentiation, growth, tissue regeneration, aging, and those that are triggered in response to injury and disease, are all mediated by a pluripotent population of stem or progenitor cells. Although these cells control the most fundamental aspects of biology, the cellular and molecular mechanisms that define them and that control their expansion and differentiation is not well understood. The colonial ascidian, B. schlosseri, provides a unique model to study chordate stem cell biology. Regeneration is a major part of the life history of Botryllus: in a highly coordinated developmental process, Botryllus adults regenerate themselves, including all somatic tissues and the germline, every week. In addition, naturally occurring fusion events between individuals cause a population of primitive stem cells to mobilize, proliferate, differentiate and sometimes completely replace the germline of the host in an event called germline cell parasitism (gcp). Conserved and important aspects of stem cell biology, such as self-renewal capacity, homing or expansion, and differentiation kinetics must underlie the ability of a stem cell of one genotype to out compete a stem cell of another genotype. The overall objectives of this proposal are to use this system to prospectively isolate the cell(s) responsible for gcp, determine the cellular and molecular biological phenotype of these cell(s), and correlate those phenotypes with parasitic capability. We propose the following specific aims to accomplish the goals set forth in this study: Aim 1. To isolate and characterize the cells responsible for germline stem cell parasitism (gcp). Aim 2. To test functions of candidate stem cell markers during asexual regeneration and germline stem cell parasitism. Aim 3. To determine the ontogeny of cells responsible for germline stem cell parasitism Relevance: Characterization of stem cells in this unique chordate ancestor will help answer basic biological questions with applications in regenerative medicine.